Obesity increases the risk of post-menopausal estrogen receptor (ER)-positive breast cancer by over 50%, a significant observation given that 40% of the US population are clinically obese and that obesity is rising most rapidly in women over 60 years of age. Some of this risk can be attributed to increased estrogen production in adipose tissue and to obesity-associated increases in inflammatory cytokines, IGF-1, and circulating insulin. Recently, however, altered cholesterol metabolism, a comorbidity of obesity, has emerged as an additional independent risk factor for breast cancer in post-menopausal women. It is significant, therefore, that we demonstrated that 27-hydroxycholesterol (27HC), a primary metabolite of cholesterol, functions as an estrogen in cellular and animal models of ER-positive breast cancer. Notably, the impact of hypercholesterolemia on tumor pathology was reduced in mice in which CYP27A1, the enzyme responsible for converting cholesterol to 27HC, was ablated. The clinical relevance of these findings was confirmed in our studies demonstrating that increased expression of CYP27A1 within tumors was associated with a higher grade. This was reinforced by the findings of another group indicating that 27HC levels were significantly elevated in breast tumor tissue. We also demonstrated in animal models that elevation of 27HC dramatically increased tumor metastasis through its actions on the nuclear receptor LXR. Together, these findings establish 27HC as an important biochemical link between hypercholesterolemia and breast cancer and implicate ER and LXR as the primary mediators of these activities. Building on these findings, we hypothesize that the oxysterol, 27HC, is an endocrine/paracrine modulator of processes that impact breast tumor growth and metastasis. It is further proposed that LXR and ER are the primary mediators of the pathological actions of 27HC and that these activities result from a dysregulation of pathways upon which these receptors converge. To explore this hypothesis we will (a) define the temporal and spatial relationship between 27HC production and the development and progression of breast cancer, (b) explore therapeutic strategies to mitigate the impact of hypercholesterolemia on cancer pathology, and (c) define the pathways of pathological importance upon which ER and LXR converge and how this influences the pharmacology of 27HC. Together, these integrated experiments probe fundamental aspects of 27HC endocrinology/pharmacology that will enable a definition of the roles of ER/LXR crosstalk in breast cancer pathology. The heavy focus on development of therapeutics reflects the expectation that these studies will yield the data required to inform near term clinical trials of agents to mitigate the impact of hypercholesterolemia on breast cancer.